Efficacy of Plectranthus amboinicus (Lour.) Spreng in a Murine Model of Methicillin-Resistant Staphylococcus aureus Skin Abscesses

Abstract

The present work aimed to evaluate the effectiveness of Plectranthus amboinicus (Lour.) Spreng against MRSA clinical isolates. The in vitro antimicrobial activity of the hydroalcoholic extract (HE), the ethyl acetate (EA) fraction and its subfractions were determined by broth microdilution and bioautography against MRSA clinical isolates. The microdilution checkerboard method was used to assess in vitro drug combination studies. To induce abscess formation, bacterial suspensions were added to Citodex and inoculated subcutaneously into male Swiss mice. The treatment protocol consisted of 2 doses of HE, the EA fraction or vancomycin introduced intraperitoneally into mice 3 and 12 h after infection. The EA fraction and its subfractions presented the lowest minimal inhibitory concentrations (MIC, 0.25 to 0.5 mg/mL). The plant samples were bacteriostatic at 2x and 4x MIC and bactericidal at 100 mg/mL. The EA fraction presented synergism with vancomycin and an additive effect with ciprofloxacin. A significant reduction of abscess volume, bacterial cell counts in abscess slurries, and inflammatory scores was observed in the HE and EA fraction-treated groups. The samples were effective in treating the animals in a dose-dependent fashion. The present study proved the effectiveness of P. amboinicus fractions against MRSA using in vitro and in vivo assays.

Figure 1

Bacteriostatic and bactericidal activities of the hydroalcoholic (HE) extract (a) and the ethyl acetate fraction F4 (b) from P. amboinicus against a standard MRSA strain (ATCC 65398). The results are expressed as log 10 colony-forming units/mL.

Figure 2

Bioautography of the ethyl acetate F4 subfractions from P. amboinicus. Two thin-layer chromatography plates were tested as follows. (a) A developed thin-layer chromatography (TLC) plate was dipped into a Petri dish containing Mueller-Hinton agar previously inoculated with 10⁶ CFU/mL of MRSA. After incubation, the plate was stained with thiazolyl blue tetrazolium MTT stain. (b) A developed TLC plate was directly stained with vanillin/perchloric acid. The arrows indicate the zones of bacterial growth inhibition. The solvent solution used for elution was hexane: ethyl acetate (6 : 4).

Figure 3

Volume of subcutaneous abscesses in mice infected with an MRSA clinical isolate strain (#08), 72 h after intraperitoneal treatment with 2 doses of ethyl acetate fraction F4 from P. amboinicus at 500 mg/kg/dose. (a) Untreated animal. (b) Animal treated with fraction F4.